Macrophages are known to express, dependent on their state of development, several receptors which are specific for glycoproteins that contain specific sugars. These include receptors specific for sialic acid residues (Crocker, P., et al., J Exp Med (1986) 164:1862); receptors specific for galactose (Aminoff, D., et al., Proc Natl Acad Sci USA (1977) 74:1521; Schlepper-Schaffer, J., et al., Biochem Biophys Res Comm (1983) 115:551); and receptors specific for mannose residues (de May, et al., Proc Natl Acad Sci USA (1978) 75:1339). The mannose receptor is uniquely found on macrophages, and is not found on monocytes. The synthesis and processing of the macrophage receptor were described by Lennartz, M. R. et al., J Biol Chem (1989) 264:2385-2390.
The mannose receptor itself is a 170 kD glycoprotein which has been isolated from several sources. The human placental receptor has been characterized and the gene cloned and sequenced (Lennartz, M. R. et al., J Biol Chem (1987) 262:9942-9944; Taylor, M. E. et al., J Biol Chem (1990) 265:12156- 12162, the latter paper incorporated herein by reference).
A good deal is known about the behavior of the mannose receptor in internalizing ligands to which it binds. After internalizing the receptor ligand complex, the intracellular vesicles containing the complexes become acidic and dissociate the complex. The unoccupied receptor is returned to the cell surface while the ligand remains inside the cell. This cycle takes less than 15 minutes, and receptor molecules have half-lives of more than 30 hours, thus offering the capability to perform hundreds of cycles with respect to a single receptor. It has been shown that alveolar macrophages can accumulate about 50.times.10.sup.6 molecules of mannose-BSA ligand per cell per 24 hours. (Ezekowitz, R. A. B., et al., J Cell Sci (1988) Supp. 9:121).
While mannose binding receptors regardless of source in general show similar specificities--i.e., recognize glycoproteins with terminal mannose and fucose and, to some extent N-acetylglucosamine and glucose, it appears that the quantitative affinity of these receptors for various ligands depends on their cellular origin. For example, N-acetylglucosamine-BSA (bovine serum albumin) binds reasonably well to alveolar macrophage but binds poorly to human placental mannose receptor. Adding further complexity is the presence of an approximately 30 kD mannose binding protein which is secreted by liver hepatocytes. The gene for this protein has also been cloned and sequenced (Ezekowitz, R. A. B. (supra)).
It is known that BSA or HSA (human serum albumin) derivatized to mannose through lysyl residues in the structure is a potent binder to the mannose ligand (Stahl, P. et al., Cell (1980) 19:207-215). Most BSA preparations contain about 57 lysines, of which 30-40 are coupled with mannose in a standard mannose-BSA preparation. In addition, one of the known native targets for the receptor, yeast mannan, is a polymannose (.alpha.1-6) backbone with mono-, di- and trisaccharide sidechains linked .alpha.1,2 and .alpha.1,3 to the backbone. However, these materials are not ideal targeting agents for macrophage as they are inherently heterogeneous compositions and do not provide reproducible binding affinities satisfactory for pharmaceutical applications. It would therefore be helpful to have a defined composition as a high affinity ligand for the macrophage receptor.
Others have attempted to synthesize ligands containing mannosyl residues. For example, Ponpipom, M. M. et al., J Med Chem (1981) 24:1388-1395, describe mono-, di- and oligolysine backbones which are derivatized to mannosyl or fucosyl residues through short covalent linking arms. In addition, this paper reports polymerization of N-lipoyl-.beta.-D-mannopyranosylamine to result in an effective receptor binding ligand. The best of these was able to effect 50% inhibition of labeled mannosylated BSA binding to macrophage only at concentrations greater than 10 .mu.M.